A Handbook of Gene and Cell Therapy

Acknowledgments
Contents
1: Gene and Cell Therapy
	1.1	 The Concepts of Gene and Cell Therapy
	1.2	 Types of Gene Therapy
	1.3	 Gene Therapy Strategies
	1.4	 Choice of the Therapeutic Target
	1.5	 Administration Routes
	1.6	 Delivery Systems
	1.7	 Expression and Persistence of the Therapy
	1.8	 Cell Targeting
	1.9	 Immune Response to the Therapy
	1.10	 Highlights in the History of Gene and Cell Therapy
	1.11	 Current Status of Gene Therapy
	1.12	 Ethical Questions and Concerns About Gene and Cell Therapy
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
2: Non-viral Vectors for Gene Therapy
	2.1	 Physical Methods
		2.1.1	 Hydrodynamic Delivery
		2.1.2	 Microinjection
		2.1.3	 Electroporation
		2.1.4	 Nucleofection
		2.1.5	 Ultrasound and Sonoporation
		2.1.6	 Ballistic Gene Delivery/Gene Gun
		2.1.7	 Magnetofection and Magnetoporation
		2.1.8	 Microneedles
	2.2	 Chemical Systems
		2.2.1	 Polymer-Based Nanocarriers
		2.2.2	 Lipid-Based Systems
		2.2.3	 Inorganic Materials
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
3: Viral Vectors for Gene Therapy
	3.1	 Lentiviral Vectors
		3.1.1	 Replicative Cycle
		3.1.2	 From Lentivirus to Lentiviral Vectors
		3.1.3	 Additional Improvements to Lentiviral Vectors
		3.1.4	 Lentiviral Vector Production
		3.1.5	 Lentiviral Vectors in Clinical Trials
	3.2	 Gamma Retrovirus
	3.3	 Adenoviral Vectors
		3.3.1	 Replicative Cycle
		3.3.2	 From Adenovirus to Adenoviral Vectors
		3.3.3	 Adenoviral Vector Modifications
		3.3.4	 Adenoviral Vector Production
		3.3.5	 Adenoviral Vectors in Clinical Trials
	3.4	 Adeno-associated Virus (AAV)
		3.4.1	 Replicative Cycle
		3.4.2	 From AAV to AAV Vectors
		3.4.3	 AAV Modifications
		3.4.4	 AAV Production
		3.4.5	 AAV in Clinical Trials
	3.5	 Herpes Simplex Virus
		3.5.1	 Replicative Cycle
		3.5.2	 From HSV to HSV Vectors
		3.5.3	 HSV Modifications
		3.5.4	 HSV Production
		3.5.5	 HSV in Clinical Trials
	3.6	 Vaccinia
		3.6.1	 Replicative Cycle
		3.6.2	 From Vaccinia Virus to Vaccinia Vectors
		3.6.3	 Vaccinia Modifications
		3.6.4	 Vaccinia Vector Production
		3.6.5	 Vaccinia in Clinical Trials
	3.7	 Baculovirus
		3.7.1	 Replicative Cycle
		3.7.2	 From Baculovirus to Baculovirus Vectors
		3.7.3	 Baculovirus Modifications
		3.7.4	 Baculovirus Production
		3.7.5	 Baculovirus in Clinical Trials
	3.8	 Choice of the Viral Vector
	3.9	 Oncolytic Virus Applications
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
4: Barriers to Gene Delivery
	4.1	 Extracellular Barriers
		4.1.1	 Unspecific Interactions
		4.1.2	 Endothelial Barriers
		4.1.3	 Inflammatory and Immune Response
	4.2	 Intracellular Barriers
		4.2.1	 Cellular Uptake
		4.2.2	 Endosomal Escape
		4.2.3	 Intracellular Trafficking
		4.2.4	 Nuclear Delivery
	4.3	 Technical Barriers
		4.3.1	 Physical Restrictions
		4.3.2	 Cellular Targeting
		4.3.3	 Gene Persistence
		4.3.4	 Sustainable Gene Expression
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
5: Stem Cells and Tissue Regeneration
	5.1	 Adult and Fetal Neural Stem Cells
	5.2	 Embryonic Stem Cells
	5.3	 Stem Cells Generated Through Cell Reprogramming
		5.3.1	 Induced Pluripotent Stem Cells (iPSC)
		5.3.2	 Chemically Induced Pluripotent Stem Cells (CiPSC)
		5.3.3	 Epigenetic, Metabolic and Cellular Pathway Modulation by Small Molecules to Induce Pluripotency
		5.3.4	 Direct Reprogramming Mediated by Reprogramming Factors and/or Small Molecules
		5.3.5	 iPSC and iPSC-Derived Neural Progenitors’ Issues
	5.4	 Brain in a Dish: From Organoids to 3D Bioprinting
		5.4.1	 Organoids
		5.4.2	 Three-Dimensional (3D) Bioprinting
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
6: Gene Therapy Strategies: Gene Augmentation
	6.1	 Gene Replacement
	6.2	 Gene Addition
	6.3	 Gene Addition to Modulate Autophagy
		6.3.1	 The Autophagy Pathway
		6.3.2	 Autophagy Terms Glossary
		6.3.3	 Upregulation of the Autophagy Pathway as a Therapeutic Strategy for Machado-Joseph Disease/Spinocerebellar Ataxia Type 3
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
7: Gene Therapy Strategies: Gene Silencing
	7.1	 Antisense Oligonucleotides
		7.1.1	 ASOs Generations
		7.1.2	 Important Considerations for the Use of ASOs in Gene Therapy
		7.1.3	 Functional Mechanisms
		7.1.4	 ASOs Applications in Gene Therapy
	7.2	 RNA Interference
		7.2.1	 Gene Expression Regulation in Eukaryotes
		7.2.2	 The Small Interfering RNA Pathway
		7.2.3	 The MicroRNA Pathway
		7.2.4	 Small Interfering RNAs Versus MicroRNAs
		7.2.5	 Small Interfering RNAs Versus Short Hairpin RNAs
		7.2.6	 Gene Therapy Applications of RNAi
		7.2.7	 RNAi Terms Glossary
		7.2.8	 Gene Silencing as Therapy for Machado-Joseph Disease/Spinocerebellar Ataxia Type 3
	7.3	 Future Prospects on Gene Silencing
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
8: Gene Editing
	8.1	 Rewriting the Typewriter
	8.2	 The Basis of Gene Editing
	8.3	 DNA Double-Strand Break Repair Mechanisms
		8.3.1	 Nonhomologous End-Joining
		8.3.2	 Homology-Directed Repair
		8.3.3	 Manipulating DNA Double-Strand Break Repair Mechanisms to Edit Genomes
	8.4	 Programmable Nucleases Used in Gene Editing
		8.4.1	 Meganucleases
		8.4.2	 Zinc-Finger Nucleases
		8.4.3	 Transcription Activator-Like Effector Nucleases
		8.4.4	 CRISPR-Cas Systems
		8.4.5	 Comparing the Four Classes of Programmable Nucleases Used in Gene Editing
	8.5	 Editing Genes Using CRISPR-Cas
	8.6	 Expanding the Possibilities of Gene Editing with CRISPR-Cas
		8.6.1	 Gene Editing Beyond DNA Double-Strand Breaks
		8.6.2	 Cas Variants
	8.7	 Limitations and Challenges to Current Gene Editing Strategies
	8.8	 Gene Editing as a Tool for Human Disease Therapy
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
9: Gene Therapy Applications
	9.1	 Preclinical Studies and Clinical Trials
	9.2	 Gene Therapy for Cancer
		9.2.1	 Suicide Gene Therapy
		9.2.2	 Oncolytic Gene Therapy
		9.2.3	 Immunomodulatory Gene Therapy
		9.2.4	 Tumor-Suppressor Gene Therapy
		9.2.5	 Comparing Gene Therapy Strategies for Cancer
		9.2.6	 Challenges to Gene Therapy Targeting Cancer
	9.3	 Gene Therapy for Eye Conditions
		9.3.1	 A Privileged Immunologic Organ
		9.3.2	 Gene Delivery Routes and Vectors
		9.3.3	 Gene Therapy Trials for Ocular Conditions
		9.3.4	 Challenges to Gene Therapy Targeting the Eye
	9.4	 Gene Therapy for Cardiovascular Diseases (CVDs)
	9.5	 Gene Therapy for Neurodegenerative Diseases
		9.5.1	 Administration Routes to the CNS
		9.5.2	 Candidate Conditions for Gene Therapy
		9.5.3	 Vectors for Delivering Genes into the CNS
		9.5.4	 Alzheimer’s Disease
		9.5.5	 Parkinson’s Disease
		9.5.6	 Lysosomal Storage Diseases (LSDs)
		9.5.7	 Amyotrophic Lateral Sclerosis
		9.5.8	 Polyglutamine Diseases
	9.6	 Future Prospects for Gene Therapy Studies
	This Chapter in a Nutshell
	Review Questions
	References and Further Reading
Solutions to the Review Questions of Each Chapter